[14] Antibody-coated plastic tubes in radioimmunoassay

Parsons, George H.

doi:10.1016/0076-6879(81)73067-2

Cited by 30 publications

(8 citation statements)

References 15 publications

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“…It is generally known that various brands and lots of polystyrene plates differ in adsorption properties for antigens (3,4,10,15). Indeed, a common procedure when adopting ELISA to measurement of antigens or antibodies in a new system is to test several different types of plates in the hope that one may give better results than Effects of various conjugate dilutions on detection of competition between BSA and human IgG on type B plates.…”

Section: Discussionmentioning

confidence: 99%

Principles, problems, and strategies in the use of antigenic mixtures for the enzyme-linked immunosorbent assay

Kenny¹,

Dunsmoor²

1983

J Clin Microbiol

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Competition between proteins and other macromolecules for adsorption sites on plastic was studied with the enzyme-linked immunosorbent assay (ELISA) to determine effects of the use of antigenic mixtures or extracts of organisms on assays of antibodies and antigens by ELISA. A comparison of a number of different polystyrene microplates with bovine albumin and human immunoglobulin G (IgG) as antigens showed two major classes of plates: those which adsorbed albumin poorly and those which adsorbed albumin well. IgG adsorbed well on all plates, but plates which adsorbed albumin best also gave significant background levels of nonspecific binding of conjugate. When mixtures of IgG and bovine serum albumin were used as coating antigens, significant competition was observed; the component present at 1% or less in the mixture was essentially undetectable unless excessive amounts of conjugate were used. The important factor was the ratio of competitor to antigen, not the absolute amount. Other proteins (ovalbumin, rabbit albumin, human albumin, and gelatin) were equally effective competitors for adsorption sites on plastic. Nonionic detergents (Tween 20, and Triton X-100) were strong competitors even at 10:1 competitor-to-antigen ratios. In antigen capture assays, normal serum components blocked attachment of antigen-specific IgG, but this competition could be lessened to a degree by the use of strongly binding polystyrene plates. In indirect ELISA for measurement of serum antibody, the use of antigenic mixtures gave significantly lower antibody titers when the desired antigen was less than 1% of the total protein coated. Therefore, the use of mixed or crude antigens in ELISA presents significant problems concerning the sensitivity and specificity of tests. The enzyme-linked immunosorbent assay (ELISA) (9) is a major tool for detecting antigens and antibodies in a wide variety of diseases (20, 21). Although the method was initially devised for measurement of antibodies to individual proteins, a large number of assays for individual disease agents have been devised using partially purified or even unpurified antigens. Such assays are not uniformly successful, and when they are it is not possible to define the reacting antigen(s). If we were to wait until suitably purified antigens were available, a number of currently useful assays would not be available for serodiagnosis. It has been recognized that components of antigenic mixtures (i.e., an extract of a microorganism) likely compete with each other for the limited sites on plastic surfaces (4, 5, 13, 16). Thus, studies to determine the nature of competition are necessary to define conditions under which ELISAs can best be performed with partially purified or unpurified antigens. Proteins apparently attach to plastic surfaces by hydrophobic interactions (12, 19), and the

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Section: Discussionmentioning

confidence: 99%

Principles, problems, and strategies in the use of antigenic mixtures for the enzyme-linked immunosorbent assay

Kenny¹,

Dunsmoor²

1983

J Clin Microbiol

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“…Solid-phase method. The preparation of coated polystyrene plates and the reaction of these plates with '251-Px were performed essentially according to the method of Parsons (1981) and Norgaard-Pedersen et al (1983). Recombination test wells were coated with Ig purified from lung adenocarcinoma pleural effusions as described above.…”

Section: Recombination Of I'si-fi With Ig From Cancer Patientsmentioning

confidence: 99%

Characterization of a lung‐cancer‐associated auto‐antigen

Zeng

Alpert

1992

Intl Journal of Cancer

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An antigen, protein X (Px), was purified from immune complexes isolated from malignant pleural effusions from patients with adenocarcinoma of the lung by EDTA treatment, PEG 8000 precipitation, protein A affinity chromatography, and Sephadex G-200 separation in the presence of 3 M NaCl. The purified antigen had a M(r) 17,000 by SDS-PAGE, and consisted of isoelectric species of pI 6.3 and 6.6. Purified Px recombined with Ig isolated from pleural fluids from patients with lung adenocarcinoma, but not with Ig from patients with breast carcinoma. Using an autologous human and heterologous chicken antibody, Px was found, by immunohistology, in the cytoplasm of some of the well-differentiated lung adenocarcinoma cells, but was not seen in normal lung or a variety of other malignant tissues. A liquid-phase competitive-inhibition RIA was developed. Over 30 ng/ml of Px were found in 9 of 15 pleural fluids from patients with lung carcinoma, none of 20 from patients with breast, ovary, stomach or colon cancer, and in 3 of 15 patients with unknown primary tumor. Our data suggest that Px may be a lung-cancer-associated autoantigen which can elicit a host humoral response in vivo.

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“…An effectively automated sandwich procedure, for the detection of Salmonella DNA in biological samples, was first described in 1985 (12). Two consecutive restriction fragments derived from the genome of S. typhimurium were isolated; one of these fragments, A, was immobilized onto polypropylene solid-phase receptacles (SPR's), which resemble pipet tips closed at the top, by well-characterized protein-DNA complexing procedures (100,101). The other fragment, B, was 32P-labelled; denatured sample was mixed with A and B simultaneously, and hybridization and washing were performed in a commercially available automated immunoassay system (102).…”

Section: Variations On the Sandwich Themementioning

confidence: 99%

Nucleic acid analysis by sandwich hybridization

Nicholls

Malcolm

1989

Clinical Laboratory Analysis

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P e t e r J o h n N i c h o l l s . A T h e s i s S u b m it t e d f o r t h e D e g r e e o f D o c t o r o f P h i lo s o p h y o f t h e U n i v e r s i t y o f L o n d o n . C h a r i n g C r o s s a n d W e s t m in s t e r M e d i c a l S c h o o l.M a r c h , 1 9 8 9 . 2 A b s t r a c t . T h e n u c l e i c a c i d -b a s e d t e s t s c u r r e n t l y u s e d f o r t h e d e t e c t io n o f i n h e r i t e d a n d i n f e c t i o u s d i s e a s e a r e s e n s i t i v e a n d s p e c i f i c , b u t n o t a lw a y s r e p r o d u c ib le , b e c a u s e o f p r o c e d u r a l c o m p l e x i t i e s ; t h i s i n t u r n m eans t h a t t h e y a r e n o t a m e n a b le t o a u t o m a t io n . S u c h t e s t s a r e n o t c o s t -e f f e c t i v e , a r e l i m i t e d i n t h e num ber o f s a m p le s w h ic h c a n b e p r o c e s s e d a t o n c e , a n d m o s t com m only r e l y o n u n s t a b le i s o t o p i c a l l y l a b e l l e d p r o b e s , w h ic h a r e h a z a r d o u s t o h a n d le a n d d i s p o s e o f . A n u c l e i c a c i d s a n d w ic h h y b r i d i z a t i o n a s s a y h a s b e e n d e v e lo p e d , i n i t i a l l y u s i n g c lo n e d hum an p a p i l l o m a v i r u s (H PV) s e q u e n c e s a s a m o d e l, a n d s u b s e q u e n t ly f o r t h e d e t e c t io n a n d t y p i n g o f H PV i n f e c t i o n i n c e r v i c a l s c r a p e s . T h e i m p l i c a t i o n s o f c e r v i c a l HPV i n f e c t i o n a r e o f c l i n i c a l s i g n i f i c a n c e , b e c a u s e o f t h e r e l a t i v e l y r e c e n t a c c u m u la t io n o f e v id e n c e l i n k i n g s p e c i f i c H PV t y p e s w i t h t h e a e t i o l o g y o f c e r v i c a l c a n c e r . C o n s i d e r i n g t h a t v i r a l t y p e c o u ld h a v e a g r e a t i n f l u e n c e o n t h e p r o g n o s i s o f i n f e c t i o n , a n d t h a t im m u n o lo g ic a l a s s a y s a r e n o t c a p a b leo f d i f f e r e n t i a t i n g b e tw e e n t y p e s , t h e s a n d w ic h h y b r i d i z a t i o n t e c h n iq u e , w h ic h c o u ld h a n d le l a r g e n um bers o f s a m p le s , w o u ld b e a n i n v a l u a b l e t o o l f o r im p r o v in g t h e e f f i c a c y o f t h e c e r v i c a l s c r e e n i n g p rogram m e. T o Mun a n d D ad, w ith a l l n y lo v e . T h e a s s a y r e q u i r e s tw o d i s t i n c t , n o n -o v e r la p p i n g ENA fr a g m e n t s , d e r iv e d fr o m a d j a c e n t p o s i t i o n s o n t h e v i r a l genom e. One f r a g m e n t (A ) i s c o v a l e n t l y a t t a c h e d t o a S-T h a n k s f o r m a k in g t h i s p o s s i b l e - 5T h e kncw n i s f i n i t e , t h e unknow n i n f i n i t e ; i n t e l l e c t u a l l y we s t a n d o n a n i s l e t i n t h e m id s t o f a n i l l i m i t a b l e o c e a n o f i n e x p l i c a b i l i t y . O u r b u s i n e s s i n e v e r y g e n e r a t io n i s t o r e c l a i m a l i t t l e m ore la n d .T .H . H u x le y , 1 8 8 7 . F i r s t a n d f o r e m o s t ,...

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[14] Antibody-coated plastic tubes in radioimmunoassay

Cited by 30 publications

References 15 publications

Principles, problems, and strategies in the use of antigenic mixtures for the enzyme-linked immunosorbent assay

Principles, problems, and strategies in the use of antigenic mixtures for the enzyme-linked immunosorbent assay

Characterization of a lung‐cancer‐associated auto‐antigen

Nucleic acid analysis by sandwich hybridization

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